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Gene Therapy for Sickle Cell Disease
Sickle-Cell Disease Sickle-cell disease is a genetic disorder of the blood that can be characterized by an abnormality in the hemoglobin molecule in red blood cells. Hemoglobin, typically composed of two alpha subunits and two beta subunits, is what allows red blood cells to carry oxygen from the lungs to all other parts of the body. A substitutional mutation of thymine for adenine in the HBB gene in chromosome 11 interferes with the coding process for the beta subunit of the hemoglobin protein. As a result, various types of beta subunits, such as hemoglobin S and hemoglobin C are produced (1). When paired with the alpha units, they cause the hemoglobin molecules to polymerize, causing the red blood cells to form a sickle shape through changes in solubility and oxygen tension (2). This renders them rigid and sticky, making it harder for them to travel through the bloodstream as well as unable to carry oxygen throughout the body (1). This can cause an array of both chronic and acute health problems, the most common being sickle cell anemia, in which patients experience a general range of symptoms including dizziness, fatigue, headache, and pale skin. The sickle cell gene is passed through an autosomal recessive inheritance pattern, meaning that both the mother and father must pass on the mutated gene in order for the offspring to be affected (3). Gene Therapy for Sickle Cell Disease Stem Cells One genetically effective therapeutic measure toward combatting sickle cell disease involves the transplantation of hematopoietic stem cells. These stem cells, known as CD34+ bone marrow cells, can even be from the affected patients themselves. A sample of bone marrow is harvested which then allows for isolation of these stem cells. Scientists then transduce a normal beta-globin gene to these cells before transplanting them back into the patient. This is performed with the help of a lentiviral vector, used to carry the HBB gene along with the isolated CD34+ bone marrow cells. Specifically, this lentiviral vector expresses a BAS3 globin gene, associated with anti-sickling properties. The addition of the BAS3 gene prevents polymerization of hemoglobin molecules in patients with sickle cell disease. With the addition of this single gene, the cells can then be injected back into the patient and will again be able to form correctly with no sign of sickling. All patients displayed a healthy, improved production of biconcave red blood cells (4). TALENs Transcription activator-like effector nucleases, or TALENs, are artificial restriction enzymes made through the fusion of a transcription activator-like effector DNA binding domain to a DNA cleavage domain in order to cut strands of DNA at specific sequences. Transcription activator-like effectors are easily engineered to bind to nearly any desired sequence of DNA. Once introduced into cells, the TALENs can be used for in situ genome editing (5). More specifically, scientists have been utilizing TALENs to correct the mutation of the HBB gene in sickle-cell disease patients. To do so, disease-specific patient-derived human induced pluripotent stem cells, or hiPSCs, are derived from sickle-cell disease patients. TALENs engineered to recognize and cleave at the HBB gene are then inserted into these hiPSCs along with a piggyBac transposon vector plasmid containing a wild type HBB gene. Once these stem cells are introduced back into the human host, they remain corrected and are able to retain full pluripotency along with a normal karyotype (6). References 1. U.S. National Library of Medicine. "Sickle cell disease." 2014. 2. Maakaron J. et al. "Sickle Cell Anemia." Medscape. 2014. 3. Mayo Clinic. "Sickle cell anemia." 2014. 4. Romero Z. et al. "β-globin gene transfer to human bone marrow for sickle cell disease." PubMed. 2013. 5. Wikipedia. "Transcription activator-like effector nucleases." 2014. 6. Sun N and Zhao H. "Seamless correction of the sickle cell disease mutation of the HBB gene in human induced pluripotent stem cells using TALENs." PubMed. 2014.